Project Summary The fidelity of mRNA translation is essential to maintain cellular homeostasis and prevent the production of aberrant proteins that could lead to disease development. Ribosomes are key components of the core translational machinery whose activity can be modulated by diverse types of nucleotide methylation on the ribosomal RNA (rRNA). A growing body of evidence indicates that the pattern of rRNA methylation is altered in cancers, leading to the synthesis of specialized ribosomes with the unique ability to translate mRNAs coding for oncogenic proteins. rRNA methylation has been shown to dynamically regulate ribosome function and influence their efficiency, accuracy and affinity for certain type of mRNAs. Hence, pathways regulating rRNA methylation can selectively drive the translation of a proteome that supports cellular transformation and tumor development. Our overarching goal is to identify pathways and key players involved in regulating the aberrant methylation of rRNA in cancers as they represent unexploited therapeutics targets. We recently uncovered that the oncogene Pelp1 is an important regulator of rRNA methylation in cancer cells. Our experimental approach, supported by preliminary data, is to elucidate the mechanisms by which Pelp1 regulates rRNA methylation and demonstrate the importance of this process for cancer progression. We hypothesize that Pelp1 supports tumorigenesis by modulating ribosomes translational activity through its ability to regulate rRNA methylation. In Aim1 we will determine whether Pelp1 regulates rRNA methylation by recruiting RNA methyltransferases to the nascent rRNA transcript. Because rRNA methylation plays an important role in regulating the translational capacity of ribosomes, we will assess whether overexpression of Pelp1 changes ribosome efficiency, fidelity, and affinity for certain types of mRNAs using translation reporter assays and a polysome profiling-sequencing approach (Aim 2). Pelp1 is overexpressed in 60-80% of breast cancers, where its level of expression directly correlates with tumor grade, metastasis, and endocrine therapy resistance. In Aim 3, we will use orthotopic mouse model of breast cancer to assess whether the ability of Pelp1 to regulate rRNA methylation is required to support tumorigenesis. Our findings suggest that by regulating rRNA methylation, Pelp1 participates in the translational reprograming of cancer cells, thereby promoting the selective translation of oncogenic genes. The successful completion of our proposed study will yield important insight into mechanisms of aberrant mRNA translation and it roles in oncogenesis.